Compositions for Nanoemulsion Delivery Systems

ABSTRACT

The present disclosure provides a pharmaceutically acceptable, stable, and optically clear oil-in-water nanoemulsions (intensity-averaged diameter&lt;100 nm) with an oil phase comprising &gt;10% w/v of long chain triglyceride, total surfactant and cosurfactant concentration less than that of oil phase and without the use of alcohol as cosolvent in the aqueous phase. The nanoemulsions of this disclosure have extremely favorable particle size distribution, optical clarity, and product stability against Ostwald ripening with high levels of oil concentrations. Poorly water soluble, therapeutically active agents and others can be incorporated in the nanoemulsion systems to improve their solubility/stability in aqueous medium or to enhance their delivery for use in pharmaceutical, food, cosmetic, and other applications by oral, intravenous, subcutaneous, intra muscular, inhalation, nasal, topical, ocular, and transdermal routes.

PRIORITY

This application claims priority to U.S. utility application Ser. No.16/298,369, filed on Mar. 11, 2019, which claims priority to U.S.utility application Ser. No. 14/623,150 filed on Feb. 16, 2015, whichclaims priority to U.S. provisional application No. 61/939,965 filed onFeb. 14, 2014, the contents of all of which are fully incorporated byreference its entirety.

FIELD OF INVENTION

The present invention relates to compositions of stable, and opticallyclear, oil-in-water nanoemulsions with intensity-averaged diameter lessthan 100 nm, comprising an oil phase with at least 10% w/v long chaintriglyceride and eliminating the need of alcohol in the aqueous phase asa cosolvent. The processes for the said compositions and theirapplications as a carrier for active components in pharmaceutical, food,nutraceutical, and cosmetic industries are described in the invention.

BACKGROUND

Drug's low solubility constitute a very difficult challenge to develop abioavailable and physical stable pharmaceutical and nutraceuticalproduct, particularly when intravenous or oral solutions are needed. Anumber of approaches for preparing intravenous and oral liquidcompositions of sparingly or poorly water-soluble basic drugs areavailable. These methods include micellar solubilization or drugnanoparticle suspension by means of surface-active agents; formation ofcomplexes with cyclodextrin and its derivatives (Hydroxypropylbeta-Cyclodextrin (HPBCD) and sulfobutylether-β-cyclodextrin (SBECD));use of various co-solvent systems; and formation of salt with strongacid with a low solution pH. However, for micellar system, surfactantshave been implicated by adverse effects such as irritation, hemolysisand histamine reaction and severe anaphylaxis reaction, and fornanosuspension system wherein pure drug particles of nanosize stabilizedby polymer and surfactants, potential catalytic degradation of drugsubstance due to higher exposure area to aqueous media and thesurrounding surfactants has been reported; taste masking and injectionpain is another issue for the micellar/nanosuspension system due to ahigher concentration of free drug available in the aqueous medium;co-solvent systems are known for causing precipitation, injection painand phlebitis; potential nephrotoxicity and bradycardia and reduction ofblood pressure caused by cyclodextrin and its derivatives and thepotential concerns of cyclodextrin binding with co-administeredlipophilic drugs have been reported; and the low solution pH of weaklybasic salt formed with strong acid will cause drug-excipient and productstability issue and cause tasting issue, injection site irritation andpain as a result of precipitation of the drug as free base when contactwith blood at neutral pH. In summary, each of these methods listed abovehas its inherent limitations and are insufficient to formulate lowsoluble drugs in a biocompatible vehicle having sufficient stability,minimum side effects, and appropriate pharmadynamic profiles as eitherintravenous, ocular, nasal, topical, transdermal, or oraladministration.

Oil-in-water emulsions, which are made of oil droplets dispersed in anaqueous continuous phase, offers a unique system that can address drugsolubility and stability problems with many applications in productssuch as pharmaceutical, food, and cosmetics. One of the uses ofemulsions is to deliver active pharmaceutical ingredients and activecomponents for use in topical, nutraceutical, oral, nasal, ocular, andpharmaceuticals. Active components that are soluble in oil can bedissolved/dispersed within the oil phase of the emulsion, and activecomponents that are poorly soluble in both oil and water can beincorporated at the interfacial region of the emulsion as well.

Based on its appearance or particle size, emulsion can be classifiedinto three types: macroemulsion, microemulsion and nanoemulsion.Macroemulsion with average size range of >100 nm tend to have a cloudymilky appearance because the many interfaces scatter light as it passesthrough the emulsion. Microemulsions and nanoemulsions, with averagedroplet sizes below 100 nm are two special classes of emulsions,appearing optically clear (translucent or transparent). This property isdue to the fact that light are scattered by the droplets only if theirsizes exceed about one-quarter of the wavelength of the incident light.When the mean droplet size in the emulsion is below about 100 nm,preferably below 70 nm, the light can penetrate through the emulsionwithout being scattered. Microemulsions are thermodynamic stable system,spontaneously formed by “solubilizing” oil molecules with a mixture ofsurfactants, co-surfactants, and co-solvents. Whereas nanoemulsion arethermodynamic metastable system, formation of which require externalenergy to break down oil droplet to below 100 nm level.

Conventional oil in water emulsions, i.e. macroemulsion, are inherentlyunstable system and will not form spontaneously. Energy input such asmechanical mixing, homogenizing, or ultrasound is required to form amacroemulsion; and macroemulsions tend to revert back to the stablestate of the phases resulting phase separation such as agglomeration andcreaming. Besides physical instability, relative large droplet size ofmacroemulsions have a lower interfacial area to volume ratio that limitthe ability of macroemulsion to efficiently dissolve poorly solublecompounds, which are soluble either inside oil or at the oil-waterinterface; and the opacity of macroemulsion reduce visual clarity whenadministration to eyes. Furthermore, the release of the activeingredient from macroemulsions comprised of long chain triglyceride oilby oral administration may be be often limited by the rate and extent oflipolysis. The rate of triglyceride emulsion digestion in GI tract is afunction of pH, lipase concentration, bile salt and emulsion surfacearea. Emulsions with higher surface area to volume ratios would undergofaster lipolysis than those with low surface area to volume ratios.

Formulations of emulsion into average size below 100 nm are exceptionsto those disadvantages, wherein microemulsions are thermodynamicallystable, and nanoemulsions, though thermodynamically metastable innature, could still maintain its kinetic stability for a long timeperiod due to extremely small size. The formation of emulsions below 100nm has added benefits of increasing the relative amount of interfacialarea considerably. An increase in the relative amount of interfacialarea can lead to a greater ability to dissolve poorly soluble activecomponents into the aqueous medium and a faster rate of digestion bylipolysis as compared to macro emulsions and thus a faster release ofthe active ingredient from the oil droplets. Due to small size below 100nm, microemulsion or nanoemulsion has added benefit of aiding activecompounds to penetrate epithelial mucosal layers such as eyes, skin,nasal, lung, GI tract, tumor, blood vein, and blood-brain barrier.

Despite of the similarity in particle size less than 100 nm andperformance in delivery of active compounds, microemulsions andnanoemulsions are fundamentally different. Despite of its thermodynamicstability, the required surfactant concentration for a microemulsionsystem significantly exceeds the concentration of the oil phase and isnormally several times higher than that of nanoemulsion. Because of manyundesirable side-effects caused by surfactants and due to the governmentregulations of daily intake limits of many surfactants, microemulsionsare disadvantageous in many pharmaceutical applications such asintravenous, ocular, and oral administration as compared tonanoemulsions. In addition, many surfactants have a bitter taste whenpresent in the foods/dosage form, which may cause palatability issues.Furthermore, the physical stability of a microemulsion system is oftenaffected by dilution, by heating, or by changing pH levels.

A nanoemulsion—even though it will not be formed spontaneously and onlymaintain a kinetic stability—uses much less surfactant due to the aid ofmechanic shear to break down oil droplets to nanosize level in thepresence of water and surfactant. This leads to a more tolerable systemfrom a toxicological and regulatory perspective. Similarly asmicroemulsions, nanoemulsions can have the benefit of appearingtranslucent as a result of their small size. Nanoemulsions have the samehigh interfacial area to volume ratio as microemulsions, which can aidin the dissolution of poorly soluble compounds and aid in the rapiddigestion of the emulsion by lipolysis. In contrast to microemulsions,nanoemulsions maintain its physical stability upon dilution and/orchange in pH.

Despite of many advantages over macroemulsion and microemulsions,nanoemulsions have its limitations, i.e. kinetic stability—the particlesize may increase over time via Ostwald ripening. An increase innanoemulsion particle size over time is disadvantageous as thenanoemulsion will lose its clarity accompanied with a correspondingdecrease in interface surface area. In order to achieve stablenanoemulsion with average particle size below 100 nm, low viscosity oilsincluding short chain triglycerides or medium chain triglycerides suchas Migloyol are often utilized to make nanoemulsion, the disadvantage ofwhich is the tendency of Ostwald ripening due to smaller molecular size,high aqueous solubility and low viscosity of short/medium chaintriglycerides. To improve physical stability of nanoemulsion, long chaintriglycerides with very low aqueous solubility may be employed. However,it is known that the large molecular volume and high viscosity of longchain triglycerides prevents them from readily forming optically clear(transparent or translucent) nanoemulsions with a high level of oilcontent. Therefore, in order to form translucent nanoemulsion comprisinglong chain triglyceride, either high level of small molecule weightorganic cosolvent such as alcohol, or higher levels of toxic surfactantssuch as Cremophor EL relative to that of oil phase, are commonlyutilized to reduce the surface tension of the oil droplets comprisinglong chain triglyceride. This, however may lead to an intolerable systemfrom safety, toxicological, regulatory perspective. For example, sincephosphatidylcholines (egg or soy lecithin) are naturally occurringnon-toxic, biocompatible surfactants, the preparation of lecithin-basedemulsions is of considerable pharmaceutical interest. However, sincelecithin has a strong tendency to form liquid crystalline structures ata relatively low concentration, particularly in water phase, addingalcohol to the aqueous phase as a cosolvent is necessary in order toreduce the interfacial tension thus to produce lecithin-basedmicroemulsion/nanoemulsion comprising long chain triglyceride oils.However, alcohol is known to induce toxic side effects such as enzymeinduction, drug-drug interaction, or damage to central nerve system.

Therefore, challenges remain in creating nanoemulsion with its oil phasecomprising long chain triglyceride with a high level of oil content ofthe composition, wherein the emulsion has an average size of less than100 nm (intensity averaged), maintains good stability against Ostwaldripening and optical translucency, uses biocompatible surfactant andrelative low level of other surfactants, and eliminate the use ofundesirable alcohol as co-solvent in the aqueous phase. The creation ofsuch nanoemulsions would have advantages in improving emulsion productsafety, efficacy, stability, tolerability, and taste.

SUMMARY OF INVENTION

To address the above mentioned flaws and problems in the current art,there is a need in the art for an optically clear nanoemulsion systemwith its oil phase comprising at least 10% w/v of long chaintriglyceride, which has an intensity-averaged droplet size of less than100 nm and good stability against Ostwald ripening, uses biocompatiblesurfactant and low level of other surfactants (<15%), and eliminates theuse of undesirable alcohol as a co solvent in the aqueous phase.

Considering the aforementioned problems, the present disclosure providesa surprising result that aqueous-based oil/water nanoemulsioncompositions with intensity-average mean droplet size of <100 nm,unexpectedly gives extremely favorable particle size distribution,optical clarity, and product stability against Ostwald ripening with theoil phase concentration up to 50% of the composition. Poorly watersoluble, therapeutically active agents and others can be incorporatedinto the nanoemulsion systems to improve their solubility and stabilityin aqueous medium and for active component delivery.

It is an object of this invention to provide a stable optically clearoil/water nanoemulsion comprising and oil phase comprising long chaintriglycerides and/or other oils; and ionizable surfactant and cosurfactant(s); an aqueous phase containing no alcohol as a co solvent;and a pH adjusting agent.

The oil phase of the emulsion composition is at least of 0.5-50% w/v ofthe composition, comprising >10% of long chain triglyceride in the oilphase.

The ionizable surfactant is a biocompatible ionizable surfactant or itsderivative such as egg or soy lecithin in combination withpharmaceutical acceptable co-surfactant(s). The total surfactant andcosurfactant concentration is <25% w/v of the composition where theratio of surfactant to cosurfactant is in the range of 10:0.1 to 0.1:10,10:1 to 1:10, 10:1 to 1:5, or 5:1 to 1:5, and total concentration ofsurfactant and co surfactant is less than 100% w/w of the oil phase.

The aqueous phase comprises water and contains no alcohol as the cosolvent.

It is another object of this invention to provide a method to make astable, optically clear, oil/water nanoemulsion composition comprisinglong chain triglyceride with intensity-averaged oil droplets <100 nm andwithout the use of alcohol as co-solvent in the aqueous phase, saidmethod comprising the steps of: a) preparing an oil phase comprisinglong chain triglyceride; b) preparing an aqueous phase comprising waterand pH adjustment agent; c) incorporating biocompatible surfactant andco-surfactant(s) either in the oil phase or in the aqueous phase; d)dispersing the oil phase in the aqueous phase to form a coarse emulsion;e) forming a final emulsion by sonicating or high pressure homogenizingthe emulsion of step d); and f) adjusting pH.

It is yet another object of this invention is to provide a method to usethe optically clear nanoemulsion system composition in treatment of ahuman or an animal, where the composition comprises long chaintriglyceride with intensity-averaged oil droplets <100 nm and withoutusing alcohol as the organic cosolvent for therapeutic agents and othersfor use in pharmaceutical, food, cosmetic, and other applications byoral, intravenous, subcutaneous, intra muscular, inhalation, nasal,topical, ocular, and transdermal routes, with such stability and purityto meet the requirements of the applicable compendium, the Food and DrugAdministration, and GMP. Said method comprises the steps of: a)providing in a liquid form of an oil/water nanoemulsion compositionprepared by dispersing/dissolving therapeutic active agent or otheragents in an oil carrier; b) preparing an aqueous phase comprising waterand pH adjustment agent; c) dispersing the oil phase into the aqueousphase by sonicating or homogenizing to form oil droplets; and d)administering the said nanoemulsion composition to human or animals.

It is yet another object of this invention to provide a method to treata human or an animal by using a optically clear nanoemulsion systemcomposition comprising long chain triglyceride with intensity-averagedoil droplets <100 nm and without using alcohol as an organic cosolventfor therapeutic agents and others for use in pharmaceutical, food,cosmetic, and other applications by oral, intravenous, subcutaneous,intra muscular, inhalation, nasal, topical, ocular, and transdermalroutes, with such stability and purity to meet the requirements of theapplicable compendium, the Food and Drug Administration, and GMP. Themethod comprises the steps of: a) providing in a liquid form of anoil/water nanoemulsion composition prepared by dispersing the oil phaseinto the aqueous phase by sonicating or homogenizing to formnanoemulsion; b) adding therapeutic active agent or others agents intothe oil/water nanoemulsion from step a) and mixing to dissolve thetherapeutic active agent or others agents into the oil phase; and c)administering the said nanoemulsion composition to human or animals.

It is yet another object of this invention to provide a method to usethe optically clear nanoemulsion system composition for treating a humanor an animal where the composition comprises long chain triglyceridewith intensity-averaged oil droplets <100 nm and without using alcoholas an organic cosolvent for therapeutic agents and others for use inpharmaceutical, food, cosmetic, and other applications by oral,intravenous, subcutaneous, intra muscular, inhalation, nasal, topical,ocular, and transdermal routes, with such stability and purity to meetthe requirements of the applicable compendium, the Food and DrugAdministration, and GMP. The method comprises the steps of: a) providingin a liquid form of an oil/water coarse emulsion composition prepared bydispersing the oil phase into the aqueous phase to form coarse emulsion;b) Adding therapeutic active agent or others agents into the oil/watercoarse emulsion from step a), and mixing to dissolve/disperse thetherapeutic active agent or others agents into the oil phase; c)obtaining a liquid form of an oil/water nanoemulsion by sonicating orhomogenizing; and d) administering the said nanoemulsion composition tohuman or animals.

SHORT DESCRIPTION OF THE DRAWINGS

FIG. 1. Comparison of particle size distribution of nanoemulsion madewith LCT/lecithin/polysorbate 80 (Example 1) vs macroemulsion made withonly lecithin (Example 7). The average particle size of the nanoemulsionis 47 nm while the average of the macroemulsion is 177 nm.

FIG. 2. Translucent nanoemulsion from Example 5.

FIG. 3. Particle size distribution of nanoemulsion made withLCT/lecithin/polysorbate 80 (Example 1) after 5 months storage at 40°C./75% RH. The figure shows that the particle size distribution isalmost same before and after the storage.

FIG. 4. Particle size distribution of nanoemulsion made withLCT/lecithin/polysorbate 80 and cyclosporine (Example 2) after 2 and 5months storage at 40° C./75% RH. No significant change in particle sizeand optical clarity was observed. The average particle size was 38 nmfor initial and 45 nm after 5 month at 40° C.

FIG. 5. Particle size distribution of nanoemulsion made withLCT/MCT/lecithin/polysorbate 80 and cyclosporine (Example 3) after 2months storage at 40° C./75% RH.under refrigeration (˜2-8° C.)/Thefigure shows that the particle size distribution is almost same beforeand after storage.

FIG. 6. Particle size distribution of nanoemulsion made withLCT/lecithin/polysorbate 80 and cyclosporine (Example 5) after 12 monthsstorage at room temperature and refrigeration (˜2-8° C.). No significantchange in particle size and optical clarity was observed in eithercondition. The average particle size was 38 nm initially, 39 nm underrefrigeration for 12 month, and 49 nm at 25° C. for 12 months.

DETAILED DESCRIPTION OF THE INVENTION

The term of “emulsion” is defined as a system (as fat in milk)consisting of a liquid dispersed with or without an emulsifier in animmiscible liquid usually in droplets of larger than colloidal size.

The term of “oil in water emulsion” refers to an emulsion system, whichis made of oil droplets dispersed in an aqueous continuous phase. Inthis disclosure, the term of “emulsion” refers to oil in water emulsionwithout exception.

The term of “microemulsion” is defined as dispersion made of water, oil,and surfactant(s) that is an isotropic and thermodynamically stablesystem with dispersed domain diameter varying approximately from 1 to100 nm, usually 10 to 50 nm. The droplet size is the Z-average orintensity weighted average size as measured by dynamic light scattering.In this disclosure, the term of “microemulsion” refers to oil in wateremulsion without exception.

The term of “nanoemulsion” is defined as dispersion made of water, oil,and surfactant(s) that is thermodynamically metastable system withdispersed domain diameter varying approximately from 1 to 100 nm,usually 10 to 50 nm. The droplet size is the Z-average or intensityweighted average size as measured by dynamic light scattering. In thesaid invention, the term of “nanoemulsion” refers to oil in wateremulsion without exception.

The term of “medium chain triglyceride” means medium-chain (6 to 12carbons) fatty acid esters of glycerol.

The term of “long chain triglyceride” means long-chain (>12 carbons)fatty acid esters of glycerol.

The term of “surfactant” means usually organic compounds that areamphiphilic, meaning they contain both hydrophobic groups andhydrophilic groups.

The term of “ionizable surfactant” means usually organic compounds thatare amphiphilic, meaning they contain both hydrophobic groups andhydrophilic groups, and its head group can be ionized at physiologicalpH range of below 10.

The term of “cosurfactant” is a surfactant that acts in addition toanother surfactant, further reducing the surface tension of a liquid.

The term of “cosolvent” is organic solvent that acts in addition toother solvent, further reducing the surface tension of a liquid.

The term of “transparent” is the physical property of allowing light topass through the material without being scattered. It follows Snell'slaw; In other words, a transparent medium allows the transport of lightand allows for image formation.

The term of “translucent” means a super-set of transparency: it allowslight to pass through, but does not necessarily follow Snell's law; Inother words, a translucent medium allows the transport of light but notallows for image formation.

The term of “optically clear” refers to either transparent ortranslucent for this invention.

The invention provides aqueous-based oil in water nanoemulsionformulation composition with mean droplet size (intensity-average, nm)of <100 nm, comprising an oil phase, a mixture of ionizable surfactantand co-surfactant, and an aqueous liquid carrier. The formulation ifthis invention comprises:

a) an oil phase comprising long chain triglyceride and/or other oil,b) a mixture of ionizable surfactant and cosurfactantd) an aqueous phase comprising water, pH adjustment agent and withoutalcohol as cosolvent.

Optionally, the emulsion formulation may also contain activecomponent(s) for pharmaceutical, nutraceutical, food and cosmeticapplication, chelate agent, antioxidant, osmotic agent, suspendingagent, preservative, and buffering agent. In some embodiments, theformulation further comprises a solubilizing agent, a flavoring agent, asweetening agent, a viscosity inducing agent, electrolyte, anothertherapeutic agent, or a combination thereof.

Combinations of the various upper and lower limits to components of thenanoemulsion, as set forth in this disclosure, can be used to providedifferent embodiments of the invention.

According to one embodiment of this invention the nanoemulsioncomprises:

a) at least 0.5-50% w/w of oil phase, which comprise long chaintriglycerideb) ionizable surfactant of 0.01-30% w/w;d) co-surfactant of 0.01-30% w/w; andd) 50-99% w/w of water.in which the oil droplets have an intensity average size of less than100 nm and the ratio of ionizable surfactant to cosurfactant is in therange from 10:0.1 to 0.1:10, 10:1 to 1:10, 10:1 to 1:5, or 5:1 to 1:5,and the ratio of total surfactant/cosurfactant to oil is less than 1:1.

In a preferred embodiment, the oil-in-water nanoemulsion comprises0.5-50 w/v % of an oil phase comprising at least 10% w/w long chaintriglyceride in the oil phase, 0.1-30% of ionizable surfactant, 0.01-30%of cosurfactant and an aqueous phase without using alcohol as cosolventin the external phase.

Oil phases in the emulsion may be a liquid or solid fat of animals, orvegetables, or of algal or synthetic origin. Those of animal origininclude oils or fats such as fish oil, cod liver oil, blubber, lard,tallow, schmaltz, and butter fat. Those of vegetable origin include oilssuch as canola oil, castor oil, cocoa butter, coconut oil, coffee seedoil, corn oil, cotton seed oil, evening primrose oil, grapeseed oil,flax seed oil, menhaden oil, mustard seed oil, olive oil, palm oil, palmkernel oil, peanut oil, poppy seed oil, rapeseed oil, rice bran oil,safflower oil, sesame oil, soybean oil, sunflower oil, palm kernel oil,hazelnut oil, sesame oil and wheat germ oil. Those of synthetic origininclude oils such as synthetic triglycerides, fractionatedtriglycerides, modified triglycerides, hydrogenated triglycerides orpartially hydrogenated and mixtures of triglycerides are also included.

Preferably oil phases in the emulsion are a pharmaceutical-grade oil,preferably triglycerides such as, but not limited to soy bean oil,safflower seed oil, olive oil, cottonseed oil, sunflower oil, fish oil(containing the omega-3 fatty acids eicosapentaenoic acid (EPA), anddocosahexaenoic acid (DHA)), castor oil, sesame oil, peanut oil, cornoil, medium chain triglycerides (such as Miglyol 812 or 810), and shortchaim triglyceride. The oil phase may also contain surfactant and/orco-surfactant such as egg lecithin, soy lecithin, and other phosphoruslipids, propylene glycol diesters, oleic acid, or monoglycerides (suchas acetylareal monoglycerides). The oil phase may also be a mixture ofsaid ingredients thereof.

The preferred lipid phase is soy bean oil, medium chain triglycerides(MCT), olive oil, and fish oil, either alone or mixture with others.

The most preferred oil phase is soy bean oil. The preferred range of oilcarrier is 0.5-50%. The most preferred rang of oil carrier is 5-20%.

Surfactants are any pharmaceutically acceptable ionizable surfactant,preferably phospholipids extracted from egg yolk or soy bean, syntheticphosphatidyl cholines or purified phosphatidyl cholines from vegetableorigin. Hydrogenated derivatives, such as phosphatidyl cholinehydrogenated (egg) and phosphatidyl choline hydrogenated (soy) may alsobe used.

The most preferred surfactant is egg lecithin. The preferred range ofsurfactant is 0.35-30%. The most preferred range of surfactant is 1-18%.

The nanoemulsion may also contain a co-surfactant that actssynergistically with the ionizable surfactant to alter the interfacialtension permitting nanoemulsion formation.

Co-surfactants may be any pharmaceutically acceptable surfactantsincluding but not limited to non-ionic surfactants such as poloxamers(for example Poloxamer 188 and 407), poloxamines, polyoxyethylenestearates, polyoxyethylene sorbitan fatty acid esters or sorbitan fattyacid esters and ionic surfactants may also be used such as cholic acidand deoxycholic acid or surface active derivatives or salts thereof. Theco-surfactant may also be selected from the group consisting of oleicacid, sodium oleate, cholic acid, sodium cholate, deoxycholic acid,deoxysodium cholate and a mixture thereof. Alcohol is excluded for useas cosurfactant or cosolvent in the external aqueous phase. Theco-surfactant is presented in the emulsion of this disclosure in therange of 0.01-30 w/v % of the composition. The ratio of surfactant tocosurfatant is in the range of 10:0.1 to 0.1:10, 10:1 to 1:10, 10:1 to1:5, or 5:1 to 1:5.

The preferred range of aqueous phase is 50-99%.

The emulsion formulation may also contain active component(s) forpharmaceutical, nutraceutical, food and cosmetic application, chelateagent, antioxidant, osmotic agent, suspending agent, preservative, andbuffering agent. In some embodiments, the emulsion may also containsolubility enhancers (excluding alcohol), chelate agent, preservative,antioxidants, stabilizers, suspending agent, pH-adjusting agents ortonicity modifying agents, such as glycerol, polymer as suspendingagent, and sweetener, etc. Stabilizers may be pH modifying agents,anti-creaming or anti-foaming agents or agents which impart stability tothe nanoemulsion.

The amount of active component in the nanoemulsion may be 0 to 50%.

The amount of other ingredient besides the active component in thenanoemulsion may be 0.5 to 50 wt %.

Desirable emulsions are stable systems of intensity-average mean dropletsize of <100 nanometer with optically clear (transparent ortranslucent). The preferred intensity-average mean droplet size is below100 nm nanometer; the most preferred intensity-average droplet size isbelow 75 nanometer.

The preferred pH range of the emulsion after manufacturing and duringstorage is below pH 10. The pH adjustment agent can be a buffer orsodium hydroxide or other pH adjustment agents or combination thereof.

The emulsion of the invention can be prepared in the following method:For the aqueous phase, water is dispensed to a container and heated toabout 40-80° C. and pH is adjusted to 1-10. For the oil phase, oil isdispensed into another container and heated to about 40-80° C.Surfactant and co-surfactant is then added to the oil and heated toabout 40° C. to about 80° C. Optionally, surfactant/cosurfactant can beadded to the aqueous phase. The aqueous and oil phases are then mixedtogether by a high shear mixer to form a coarse emulsion. The emulsionis then sonicated or homogenized with a high pressure homogenizer or amicro-fluidizer at a pressure of about 5000-30,000 psi and a temperaturerange of about 5° C. to about 70° C. until a nanoemulsion with a desireddroplet size is obtained. The pH is adjusted with pH adjustment agentsuch as sodium hydroxide to the final pH. The samples are filtered anddispensed into containers, often with nitrogen gas overlay and cappedwith stoppers. The product can be manufactured by an aseptic process orby terminal sterilization. Preferably the dosage units are autoclaved toget sterile and stable emulsions. In one embodiment, the emulsion wasautoclaved at 121° C. for 15-20 minutes. In another embodiment, theemulsion is processed aseptically under sterile environment withoutautoclave.

One embodiment of this invention is a method to make nanoemulsion withadded active component for use in human or animal treatment, the methodcomprising the steps of: a) providing in a liquid form of an oil/waternanoemulsion composition prepared by i) Adding therapeutic active agentor others agents into the oil and mixing to dissolve the therapeuticactive agent or others agents into the oil phase; ii) dispersing the oilphase comprising active component into the aqueous phase by sonicatingor homogenizing to form nanoemulsion; and b) administering the saidnanoemulsion composition to human or animals.

One embodiment of this invention is a method to make nanoemulsion withadded active component for use in human or animal treatment, said methodcomprising the steps of: a) providing in a liquid form of an oil/waternanoemulsion composition prepared by i) dispersing the oil phase intothe aqueous phase by sonicating or homogenizing to form nanoemulsion;ii) Adding therapeutic active agent or others agents into the oil/waternanoemulsion from step a) and iii) mixing to dissolve the therapeuticactive agent or others agents into the oil phase; and b) administeringthe said nanoemulsion composition to human or animals.

Another embodiment this invention is to provide a method to makenanoemulsion with added active component for use in human or animaltreatment, said method comprising the steps of a) providing in a liquidform of an oil/water coarse emulsion composition prepared by i)dispersing the oil phase into the aqueous phase to form coarse emulsion;ii) Adding therapeutic active agent or others agents into the oil/watercoarse emulsion from step i), and iii) mixing to dissolve/disperse thetherapeutic active agent or others agents into the oil phase; iv)obtaining a liquid form of an oil/water nanoemulsion by sonicating orhomogenizing; and b) administering the said nanoemulsion composition tohuman or animals.

The invention is now described by way of non-limiting examples. Theinvention comprises combinations of the embodiments and aspects of theinvention as detailed herein. Accordingly, the invention also includescombinations and sub-combinations of the individual elements of theembodiments or aspects of the invention as described herein. Otherfeatures, advantages and embodiments of the invention will becomeapparent to those skilled in the art by the following description,accompanying examples. The disclosure herein is directed to all suchvariations and modifications to such elements and methods known to thoseskilled in the art. Furthermore, the embodiments identified andillustrated herein are for exemplary purposes only, and are not meant tobe exclusive or limited in their description of the present invention. Askilled artisan would realize that various changes and modifications maybe made without diverting from the spirit of the invention.

Example 1. Preparation of Nanoemulsion Using Soybean Oil and withoutActive Ingredient

Quantities: Formula g soy bean oil 20.0 egg lecithin 7.2 polysorbate 807.2 sodium hydroxide q.s. to pH 6-9 Water for Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water nanoemulsion is prepared as follows:

-   -   1. An aqueous phase is prepared from Water for Injections,        stirred and heated to a temperature of approximately 60° C.    -   2. The aqueous phase is passed through a 0.22 micron filter and        charged to a mixing vessel.    -   3. Separately, an oil phase is prepared from soy bean oil that        has been passed through a 0.22 micron filter, polysorbate 80,        and egg lecithin, in a vessel. The mixture is stirred at a        temperature of approximately 60° C. until all ingredients are        dissolved.    -   4. The mixture is added to the aqueous phase.    -   5. This mixture is then mixed with a high shear mixer (Polytron        PT3100) at 10,000 rpm for 5 minutes to obtain a coarse emulsion.        The emulsion pH is adjusted to 6-9.    -   6. The mixture is then homogenized with a high pressure        homogenizer (APV 2000) in the range of 5,000-30,000 psi until        reaching desired particle size.    -   7. The resulting oil-in-water nanoemulsion is cooled, pH adjust        to 6-9 if necessary, and then transferred into a filling vessel.    -   8. The emulsion is then filtered with 0.22 micron filter and        filled into containers under nitrogen.

Example 2. Preparation of Nanoemulsion of Soybean Oil and Loaded withActive Drug-Cyclosporine

Quantities: Formula g cyclosporine 0.2 soy bean oil 20.0 egg lecithin7.2 polysorbate 80 7.2 glycerol 2.25 sodium hydroxide q.s. to pH 6-9Water for Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water nanoemulsion is prepared as follows:

-   -   1. An aqueous phase is prepared from glycerol, and Water for        Injections. This mixture is stirred and heated to a temperature        of approximately 60° C.    -   2. The aqueous phase is passed through a 0.22 micron filter and        charged to a mixing vessel.    -   3. Separately, an oil phase is prepared from soy bean oil that        has been passed through a 0.22 micron filter, cyclosporine,        polysorbate 80, and egg lecithin, in a vessel. The mixture is        stirred at a temperature of approximately 60° C. until all        ingredients are dissolved.    -   4. The mixture is added to the aqueous phase.    -   5. This mixture is then mixed with a high shear mixer (Polytron        PT3100) at 10,000 rpm for 5 minutes to obtain a coarse emulsion.        The emulsion pH is adjusted to 6-9.    -   6. The mixture is then homogenized with a high pressure        homogenizer (APV 2000) in the range of 5,000-30,000 psi until        desired particle size is reached.    -   7. The resulting oil-in-water nanoemulsion is cooled, pH adjust        to 6-9 if necessary, and then transferred into a filling vessel.    -   8. The emulsion is then filtered with 0.22 micron filter and        filled into containers under nitrogen.

Keratoconjunctivitis sicca (KCS) or dry eye syndrome is an eye diseasecaused by eye dryness, which, in turn, is caused by either decreasedtear production or increased tear film evaporation. It is found inhumans and some animals. KCS is the most common eye disease affecting5-6% of the population. Prevalence rises to 6-9.8% in postmenopausalwomen, and as high as 34% in the elderly Inflammation occurring inresponse to tears film hypertonicity can be suppressed by topicalimmunosuppressants, such as cyclosporine. Accordingly, the formulationssuch as described in this example and below may be used to increase tearproduction in patients whose tear production is presumed to besuppressed due to ocular inflammation associated withkeratoconjunctivitis sicca.

Example 3. Preparation of Nanoemulsion of Soybean Oil/Medium ChainTriglyceride and Loaded with Active Drug-Cyclosporine

Quantities: Formula g cyclosporine 0.2 soy bean oil 10.0 Medium chaintriglyceride 10.0 egg lecithin 7.2 Polysorbate 80 7.2 glycerol 2.25sodium hydroxide q.s. to pH 6-9 Water for Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water nanoemulsion is prepared as follows:

-   -   1. An aqueous phase is prepared from glycerol, and Water for        Injections. This mixture is stirred and heated to a temperature        of approximately 60° C.    -   2. The aqueous phase is passed through a 0.22 micron filter and        charged to a mixing vessel.    -   3. Separately, an oil phase is prepared from soy bean oil and        medium chain triglyceride (Migloyol 812) that has been passed        through a 0.22 micron filter, cyclosporine, polysorbate 80, and        egg lecithin, in a vessel. The mixture is stirred at a        temperature of approximately 60° C. until all ingredients are        dissolved.    -   4. The mixture is added to the aqueous phase.    -   5. This mixture is then mixed with a high shear mixer (Polytron        PT3100) at 10,000 rpm for 5 minutes to obtain a coarse emulsion.        The emulsion pH is adjusted to 6-9 if necessary.    -   6. The mixture is then homogenized with a high pressure        homogenizer (APV 2000) in range of 5,000-30,000 psi until        reaching desired particle size.    -   7. The resulting oil-in-water nanoemulsion is cooled, pH adjust        to 6-9 if necessary, and then transferred into a filling vessel.    -   8. The emulsion is then filtered with 0.22 micron filter and        filled into containers under nitrogen.

Example 4. Preparation of Nanoemulsion by Dilution from Example 1

Quantities: Formula g soy bean oil 5.0 egg lecithin 1.8 polysorbate 801.8 sodium hydroxide q.s. to pH 6-9 Water for Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water nanoemulsion is prepared as follows:

-   -   1. Obtain nanoemulsion sample from Example 1    -   2. Dilute the emulsion with purified water at the ratio of 1:3        v/v and mix well    -   3. Adjust the dilution pH to 6-9 if necessary and mix well    -   4. The emulsion is then filtered with 0.22 micron filter and        filled into containers under nitrogen.

Example 5. Preparation of Nanoemulsion by Dilution from Example 2

Quantities: Formula g cyclosporine 0.05 soy bean oil 5 polysorbate 801.8 egg lecithin 1.8 glycerol 2.25 sodium hydroxide q.s. to pH 6-9 Waterfor Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water nanoemulsion is prepared as follows:

-   -   1. Obtain nanoemulsion sample from Example 2    -   2. Dilute the emulsion with water solution containing 2.25%        glycerol at the ratio of 1:3 v/v and mix well    -   3. Adjust the dilution pH to 6-9 if necessary and mix well    -   4. The emulsion is then filtered with 0.22 micron filter and        filled into containers under nitrogen.

Example 6. Preparation of Nanoemulsion by Dilution from Example 3

Quantities: Formula g cyclosporine 0.05 soy bean oil 2.5 Medium chaintriglyceride 2.5 polysorbate 80 1.8 egg lecithin 1.8 glycerol 2.25sodium hydroxide q.s. to pH 6-9 Water for Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water nanoemulsion is prepared as follows:

-   -   1. Obtain nanoemulsion sample from Example 3    -   2. Dilute the emulsion with purified water containing 2.25%        glycerol at the ratio of 1:3 v/v and mix well    -   3. Adjust the dilution pH to 6-9 if necessary and mix well    -   4. The emulsion is then filtered with 0.22 micron filter and        filled into containers under nitrogen.

Example 7. Preparation of Emulsion Using Soybean Oil and Lecithin(Comparative Example)

Quantities: Formula g soy bean oil 20.0 egg lecithin 12 glycerol 2.25sodium hydroxide q.s. to pH 6-9 Water for Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water emulsion is prepared as follows:

-   -   1. An aqueous phase is prepared from glycerol, and Water for        Injections. This mixture is stirred and heated to a temperature        of approximately 60° C.    -   2. The aqueous phase is passed through a 0.22 micron filter and        charged to a mixing vessel.    -   3. Separately, an oil phase is prepared from soy bean oil that        has been passed through a 0.22 micron filter and egg lecithin,        in a vessel. The mixture is stirred at a temperature of        approximately 60° C. until all ingredients are dissolved.    -   4. The mixture is added to the aqueous phase.    -   5. This mixture is then mixed with a high shear mixer (Polytron        PT3100) at 10,000 rpm for 5 minutes to obtain a coarse emulsion.        The emulsion pH is adjusted to 6-9.    -   6. The mixture is then homogenized with a high pressure        homogenizer (APV 2000) in the range of 5,000-30,000 psi until no        more reduction in particle size.    -   7. The resulting oil-in-water nanoemulsion is cooled, pH adjust        to 6-9 if necessary, and then transferred into a filling vessel.    -   8. The emulsion is then filtered with 0.45 micron filter and        filled into containers under nitrogen.

Example 8. Preparation of Emulsion Using Soybean Oil and Lecithin andPolysorbate 80

Quantities: Formula g soy bean oil 20 egg lecithin 12 Polysorbate 80 2.4sodium hydroxide q.s. to pH 6-9 Water for Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water emulsion is prepared following that ofExample 1. The intensity averaged mean particle size of the emulsionmade by the said invention is 68 nm (D50) by dynamic light scatteringmeter

Example 9. Preparation of Emulsion Using Soybean Oil and Lecithin andPolysorbate 80

Quantities: Formula g soy bean oil 35.0 egg lecithin 2.52 Polysorbate 8012.6 sodium hydroxide q.s. to pH 6-9 Water for Injections to 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water emulsion is prepared following that ofExample 1. The intensity averaged mean particle size of the emulsionmade by the said invention is 99 nm (D50) by dynamic light scatteringmeter

Example 10. Preparation of Emulsion Using Soybean Oil and Lecithin andPoloxamer F68

Quantities: Formula g soy bean oil 13.3 egg lecithin 4.8 Poloxamer F684.8 Glycerol 6.7 sodium hydroxide q.s. to pH 6-9 Water for Injections to100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water emulsion is prepared following that ofExample 1. The intensity averaged mean particle size of the emulsionmade by the said invention is 67 nm (D50) by dynamic light scatteringmeter

Example 11. Preparation of Emulsion Using Soybean Oil and Lecithin andPolysorbate 80

Quantities: Formula g soy bean oil 5.0 egg lecithin 3.0 Polysorbate 800.6 glycerol 2.25 sodium hydroxide q.s. to pH 6-9 Water for Injectionsto 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water emulsion is prepared by diluting nanoemulsion byExample 8 with glycerol solution. The intensity averaged mean particlesize of the emulsion made by the said invention is 67 nm (D50) bydynamic light scattering meter.

Example 12. Preparation of Emulsion Using Soybean Oil and Lecithin andPoloxamer F68

Quantities: Formula g soy bean oil 20.0 egg lecithin 7.2 Poloxamer F681.27 Glycerol 10 sodium hydroxide q.s. to pH 6-9 Water for Injections to100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water emulsion is prepared following that ofExample 1. The intensity averaged mean particle size of the emulsionmade by the said invention is 89 nm (D50) by dynamic light scatteringmeter

Example 13. Preparation of Emulsion Using Soybean Oil and Lecithin andPoloxamer F68

Quantities: Formula g soy bean oil 20.0 egg lecithin 7.2 Poloxamer F68 4Propylene glycol 10 sodium hydroxide q.s. to pH 6-9 Water for Injectionsto 100 g

All processing stages are carried out under nitrogen.

An aqueous oil-in-water emulsion is prepared following that ofExample 1. The intensity averaged mean particle size of the emulsionmade by the said invention is 75 nm (D50) by dynamic light scatteringmeter

Example 14. Characterization of Nanoemulsion Size Distribution byMalvern Nano ZetaSizer

The particle size distribution of the emulsion prepared with a singleegg lecithin surfactant (Example 7) was compared again the emulsionprepared by the said invention (Example 1) (FIG. 1). The intensityaveraged mean particle size of the emulsion made by the said inventionis ˜47 nm (D50), which is optically clear and has translucent appearance(FIG. 2), whereas, the one made with the soybean oil and with only egglecithin surfactant (Example 7) has milky appearance and hasintensity-averaged mean particle size of ˜177 nm (D50).

Example 15. Stability of Nanoemulsion Made with LCT/Lecithin andPolysorbate 80

The particle size distribution of the emulsion prepared withlecithin/polysorbate 80 combination (Example 1) was monitored afterstored at 40° C./75% RH for 5 months (FIG. 3). No change in particlesize was observed after stability. The emulsion also maintainedoptically translucency after stability storage.

Example 16. Stability of Nanoemulsion Made with LCT/Lecithin andPolysorbate 80 and Loaded with Active Component

The particle size distribution of the emulsion prepared withlecithin/polysorbate 80 combination and active drug cyclosporine(Example 2) was monitored after stored at 40° C./75% RH for up to 5months (FIG. 4). No significant change in particle size was observedafter stability for up to 5 months. The average size is 38 nm forinitial and 45 nm after 5 month at 40° C. The emulsion also maintainedoptically translucency and chemical stability of cyclosporine afterstability storage.

Initial 3 months 5 months Assay (mg/mL) 1.7 1.7 1.8 D50 (nm) 38 37 45

Example 17. Stability of Nanoemulsion Made with Oil Mixture, Lecithin,and Polysorbate 80 and Loaded with Active Component

The particle size distribution of the emulsion prepared withLCT/MCT/lecithin/polysorbate 80 combination and active drug cyclosporine(Example 3) was monitored after stored at 40° C./75% RH and 2-8° C. for2 months (FIG. 5). No change in particle size was observed afterstability. The emulsion also maintained optically translucency afterstability storage.

Example 18. Stability of Nanoemulsion Made with LCT Oil, Lecithin, andPolysorbate 80 and Loaded with Active Component

The particle size distribution of the emulsion prepared withLCT/lecithin/polysorbate 80 combination and active drug cyclosporine(Example 5) was monitored after stored at controlled room temperature(˜25° C.) and under refrigeration (˜2-8° C.) for up to 14 months (FIG.6). All mean particle size was observed to be below 50 nm afterstability. The emulsion also maintained optically translucency andchemical stability for cyclosporine after stability storage.

Initial 14 month @5° C. 14 months @25° C. Assay (mg/mL) 0.45 — 0.48 D50(nm) 38 39 49

What is claimed is:
 1. An oil-in-water nanoemulsion compositioncomprising: an oil phase comprising at least 10% (w/v) long chaintriglycerides, an ionizable surfactant, and a co-surfactant, wherein atotal amount of the ionizable surfactant and the co-surfactant is about10% (w/v) to about 15% (w/v) of the composition; an aqueous phase; a pHadjusting agent, and an active component, wherein the compositioncontains no alcohol; wherein the composition demonstrates stability forat least two months at 40° C./75% RH; and wherein the mean droplet size(intensity-average, nm) of the composition is below 100 nm.
 2. Thecomposition of claim 1, wherein the composition further comprises one ormore agents chosen from a chelate agent, an antioxidant, an osmoticagent, a preservative, a suspending agent, or a buffering agent.
 3. Thecomposition of claim 1, wherein the mean droplet size(intensity-average, nm) is below 75 nm.
 4. The composition of claim 1,wherein the composition contains 10-50% w/v oil phase.
 5. Thecomposition of claim 1, wherein the nanoemulsion is stable and opticallyclear with an oil phase concentration between 10-50% w/v.
 6. Thecomposition of claim 1, wherein an oil of the oil phase is chosen fromsoybean oil, safflower seed oil, olive oil, cottonseed oil, sunfloweroil, fish oil, castor oil, sesame oil, peanut oil, corn oil, mediumchain triglycerides, or mixtures thereof.
 7. The composition of claim 1,wherein the total weight of the surfactant and the co-surfactant is lessthan or equal to that of the oil of the oil phase.
 8. The composition ofclaim 1, wherein the ratio of the surfactant and the co-surfactant is inthe range of 10:0.1 to 0.1:10, 10:1 to 1:10, 10:1 to 1:5, or 5:1 to 1:5.9. The composition of claim 1, wherein the ionizable surfactant isionizable in the pH range of 0-10.
 10. The composition of claim 1,wherein the ionizable surfactant is chosen from pharmaceuticallyacceptable biocompatible surfactants comprising phospholipids extractedfrom egg yolk or soybean, phospholipids derivatives, syntheticphosphatidyl cholines, purified phosphatidyl cholines from vegetableorigin, or hydrogenated phospholipid derivatives.
 11. The composition ofclaim 1, wherein the co-surfactant is chosen from pharmaceuticallyacceptable surfactants, comprising poloxamers, poloxamines;polyoxyethylene stearates, polyoxyethylene sorbitan fatty acid esters orsorbitan fatty acid esters; ionic surfactants including cholic acid anddeoxycholic acid; surface active derivatives or salts; oleic acid,sodium oleate, cholic acid, sodium cholate, deoxycholic acid,deoxysodium cholate or a mixture thereof.
 12. The composition of claim1, wherein the composition comprises 0.01-15% (w/v) of egg lecithin and0.01-15% (w/v) polysorbate
 80. 13. A stable, optically clear,oil-in-water nanoemulsion composition comprising: an oil phasecomprising long chain triglyceride and/or other oils, an ionizablesurfactant, and a co-surfactant, wherein the composition comprises 10%to 50% (w/v) of the oil phase, wherein the oil phase contains at least10% (w/v) of the long chain triglyceride, wherein the ionizablesurfactant comprises 0.01% to 15% (w/v) of the composition, and whereinthe co-surfactant comprises 0.01% to 15% (w/v) of the composition,wherein a total amount of the ionizable surfactant and the co-surfactantis about 10% (w/v) to about 15% (w/v) of the composition, and whereinthe ionizable surfactant and the co-surfactant are incorporated into theoil phase before the oil phase is added to the aqueous phase; an aqueousphase containing no alcohol as the cosolvent; a pH adjusting agent; andan active component other than cyclosporine; wherein the mean dropletsize (intensity-average, nm) of the composition is below 100 nm, whereinsaid droplets demonstrate stability for at least two months at 40°C./75% RH; and wherein the composition contains no alcohol.
 14. A methodfor making a stable, optically clear, oil-in-water nanoemulsioncomposition of claim 1 comprising the steps of: a) preparing an oilphase comprising a long chain triglyceride; b) preparing an aqueousphase; c) incorporating an active component other than cyclosporineeither in the oil phase or in the aqueous phase; d) incorporating abiocompatible surfactant and co-surfactant in the oil phase; e)dispersing the oil phase in the aqueous phase to form a coarse emulsion;f) forming a final emulsion by sonicating, high pressure homogenizing,or high shear processing the emulsion of step e); and adjusting the pH,wherein the composition contains no alcohol.
 15. The method of claim 14,wherein the aqueous phase of step b) includes a pH adjustment agent, andthe active component is added into the final emulsion formed in step f).16. A method for treating a subject in need therefore comprisingadministering a formulation comprising the nanoemulsion composition ofclaim 1 to a subject, wherein the active component is less than 50% w/vof the formulation.